Summary of work: Apoptosis is the most frequent morphological feature of programmed cell death thought to play a pivotal role in diverse physiological and pathological processes. These include homeostatic maintenance of tissues and organs, autoimmune diabetes and diabetic neuropathy. Farnesylation is a regulated post translational modification that allows attachement of a number of proteins, including p21Ras, to the plasma membrane. By inducing the activity of the enzyme farnesyl protein transferase, insulin increases the pool of membrane-associated p21Ras and promotes GTP loading on Ras. It has been recently documented that manumycin, a selective protein farnesylation inhibitor, blocks the antiapoptotic protection exerted by insulin in serum-starved CHO cells expressing the human insulin receptors (IR) by a mechanism that remains unclear. In this study, it was found that treatment of CHO-IR cells with manumycin blocked insulin's ability to suppress pro-apoptotic caspase-3 activity which led to time-dependent proteolytic cleavage of two nuclear target proteins. The hypothesis that inhibition of farnesylated Ras was causally related to manumycin-induced apoptosis was then tested. The results showed that the response to manumycin was found to be independent of K-Ras function because membrane association and activation of endogenous K-Ras proteins in terms of GTP loading and ERK activation were unabated following treatment with manumycin. Moreover, blocking p21Ras/Raf-1/MEK/ERK cascade by the expression of a transdominant inhibitory mSOS1 mutant in CHO-IR cells kept cells sensitive to the antiapoptotic action of insulin. In constrast, insulin dependent activation of the serine/threonine protein kinase B (also termed Akt) was blocked by 4 h treatment with manumycin, a kinetic too rapid to be explained by Ras inhibition. This study suggests that the depletion of short-lived farnesylated proteins by manumycin suppresses the antiapoptotic action of insulin at least in part by disrupting Akt activation but not that of the K-Ras/Raf-1/ERK-dependent cascade. Work is underway to identify these short-lived farnesylated proteins.